The long term goal is to develop novel therapy for liver cancer through simultaneously treating liver cancer and improving liver function, because most liver cancer patients have liver cirrhosis. A group of liver-enriched transcription factors (TFs) are master regulators of liver development and liver function, and they are tumor- suppressors down-regulated during liver carcinogenesis. Literature reports strongly suggest that restoration of the master regulator HNF4? can treat not only liver cancer, but also liver fibrosis. However, like many important oncogenes and tumor-suppressors, HNF4? is a TF that lacks known activating ligand; how to modulate undruggable TFs to treat cancer is a huge challenge. Chemicals have been developed to silence undruggable oncogenes via stabilizing a special nucleic acid structure, G-quadruplex (G4) in gene promoter. However, the lack of selectivity of G4-stabilizing chemicals toward normal cells is a bottleneck in developing G4-stabilizing chemicals as novel anticancer drugs. Our preliminary data strongly suggest that G4 motif within 5' untranslated region (UTR) is critical in suppressing protein translation of HNF4? and certain liver- enriched essential TFs. The objective of this proposal is to elucidate the importance of G4 motif within 5' UTR in translational regulation of liver-enriched TFs and cancer therapy. The central hypothesis is that G4 motifs in 5' UTR of these master regulators play a key role in suppressing their protein expression. Thus, we can increase protein expression of these master regulators using small molecules or G4 DNA oligos to destabilize G4 within 5' UTR or competitively inhibit G4-interacting proteins. This will be an exciting innovative approach to trea liver cancer and improve liver function simultaneously. This central hypothesis will be tested in 3 specific aims. Aim 1 will determine mechanism of suppression of protein expression of HNF4?1 by its 5' UTR. The working hypothesis is that formation of G4 motif within 5' UTR is essential in inhibiting protein expression. Aim 2 will determine role of G4 motif within 5' UTR of human HNF1?, HNF3?, C/EBP?, C/EBP?, HDAC3, NCOR1, and p53 in regulating protein expression. The working hypothesis is that G4 motif within 5' UTR is essential in suppressing protein expression of these TFs. Aim 3 will determine effects of G4-interacting chemicals and G4 oligos from 5' UTR of above 8 human TFs on protein expression of these TFs. The working hypothesis is that TMPYP4, polyamines, and protoporphyrin IX inhibits, whereas certain G4 DNA oligos increase protein expression of these TFs in hepatoma/hepatocytes. This study is highly novel, because it is the first to address the importance of an important nucleic acid structure, G4 in regulating the protein expression of a group of essential liver-enriched TFs. This study is highly significant, because results from this study will not only provide important novel knowledge, but also greatly aid the rational design of G4- interacting anticancer drugs, and help to develop a paradigm-shift approach to coordinately target otherwise undruggable essential liver-enriched TFs to simultaneously treat liver cancer and improve liver function. PUBLIC HEALTH RELEVANCE: The United States has increasing incidence of liver cancer, a deadly disease that is the 3rd leading cause of death from cancer worldwide. The proposed studies are of importance in the under-investigated area of how the protein expression of master regulators of liver pathophysiology is regulated by a special nucleic acid structure, G-quadruplex, which is a potential novel target for anticancer drugs. Thus, the findings are expected to enhance our understanding of liver diseases and liver cancer, and this knowledge can be used to develop novel approaches to target otherwise undruggable essential oncogenes and tumor-suppressors to treat liver cancer and liver diseases.